1. Field of the Invention
The present invention relates to a therapeutic agent for treating acute or chronic graft-versus-host disease. The present invention also relates to a therapeutic agent for treating acute or chronic graft-versus-host disease that contains mesenchymal stem cells as an active ingredient.
2. General Background and State of the Art
Graft-versus-host disease (GVHD) refers to a disease wherein the body of the patient has an immune reaction to the donor's peripheral blood or the T lymphocytes in the bone marrow that are injected during homogeneous transplantation. Namely, it is a disease that is induced by living lymphocytes which were transfused causing an immune reaction that leads to problems in liver function, skin lesions, jaundice, diarrhea, fever, pancytopenia, and so on, and in severe cases patient death.
Graft-versus-host disease can be classified largely into acute graft-versus-host disease (aGVHD) and chronic graft-versus-host disease (cGVHD). cGVHD is the most major and common side effect, occurring in 20%-70% of patients living past 100 days following blood and marrow progenitor cell transplantation, and a major cause of death following transplantation. Because cGVHD and aGVHD are not successive diseases, aGVHD requires a different approach and cGVHD is becoming the bigger problem due to developments in blood and marrow progenitor cell transplantation therapeutic methods.
cGVHD in the case of homogenous transplantations, occurs usually 4-6 months following transplantation and its occurrence within 80 days or after 1 year is uncommon. Accordingly, it can be seen that a homogeneous reaction is a major prerequisite for causing cGVHD and the pathogenesis of cGVHD goes through a long incubation period or the effect on the target organ shows up slowly. Various problems in the function of the thymus gland are being discovered in the cGVHD, and it is thought that if normal thymus glands are not removed through damage caused by treatment prior to transplantation or isoantigen/autoantigen through peripheral mechanisms, pathological graft T cells increase as a reaction, and this type of pathological CD positive T cells, as a Th2 immunological reaction, cause immune deficiencies similar to auto immune diseases including cytolytic attack, secretion of inflamed fibrosis cytokine, B cell activation, and damage to target organs through formation of auto antibodies.
Clinical symptoms of cGVHD include changes in the skin such as erythema, dryness, itchiness, pigmentation change, and maculopapular rashes; changes in hair such as thinning of hair and loss of hair, and changes in the mouth such as inflamed gums, mucositis, and lip atrophy. Aside from the various lesions appearing on the eyes, reproductive organs, liver, lungs, gastrointestinal tracts, fascia, skeletal system, serous membranes, and so forth.
While cGVHD is generally defined as GVHD that occurs after 100 days following bone marrow transplantation, the manifested conditions are more important to the diagnosis than the manifested time period. In accordance with the manifested time period of the symptoms, classification can be made between a progressive onset in which aGVHD not having been cured since occurrence shifts to cGVHD, a quiescent onset in which cGVHD appears after aGVHD has been fully cured, and de novo in which it occurs without prior appearance of aGVHD. The morbidity and death rate is highest in the progressive onset, next is quiescent onset, and it is lowest in the case of de novo. As for manifested conditions, in many cases lichenous shaped rashes in the skin and mucous coat of the mouth are the first symptoms, and while it may appear on the same parts as in aGVHD, the lesions are papulous, invasive, and covered with white scales. When compared with aGVHD from a pathological histology perspective, while legions of satellite cell necrosis can still be found, the lymphocyte infiltration shows an over consolidated band. Aside from that, the gall bladder duct is diminished, and while bile accumulation can be seen, because there may be cases in which it may be mixed with legions related to the medication or viral hepatitis, there may be cases in which it is difficult to differentiate from cGVHD.
In the case of cGVHD, because immune functions are already decreased, there is fear of serious infection during treatment, and a new effective treatment with little side effect from the treatment is keenly needed. In relation to this, many researches are being reported that state there is ability for mesenchymal stem cells to be differentiated into many organ cells and that graft-versus-hot reactions can be improved by suppressing T cells.
While mesenchymal stem cells can be propagated in an undifferentiated situation as primordial cells of the original mesoderm and be separated from various organs such as bone marrow, fat tissue, liver, tendon, synovial membrane, and umbilical cord, a single marker that can precisely define it as a mesenchymal stem cell is not in existence. However, CD14, CD34, and CD45 are well known as markers for bone marrow and SH-2(CD105), SH-3(CD73), SH-4, and Th7-1 are well known as markers for mesenchyma. Mesenchymal stem cells or mesenchymal stromal cells (MSC) express major histocompatibility complex (MHC) class 1, and MHC class can induce manifestation through interferon gamma (IFN-γ), and because it does not manifest FAS or FAS L(CD40) type costimulatory molecules, it does not induce immunological reactions, and is free from cytolysis due to cytotoxic T lymphocytes (CTL) and natural killer (NK) cells. In addition, while mesenchymal stem cells suppress proliferation of T cells through density reliance at time of mixed lymphocyte reaction (MLR) and suppress proliferation of B cells as well as formation of immunoglobulin, it is known that MHC compatibility is not a necessity for MSC immune suppression. In addition, it is known that there is no change in the activity of karyotypes or telomerases in the MSC when split 50 times.
However, because MSC that exists in the body is very rare, development of technology that isolates it is important. Currently, density gradient centrifugal separation, method using monoclonal antibodies specifically for Sca-1 or STRO-1, and separation method according to cell size are known methods for separating MSC. The inventors herein have previously developed an effective MSC separation method (Republic of Korea Public Patent No. 10-0802011) that does not require a particular mechanical device or reagent, and the above method is characterized by the fact that marrow taken from the individual is cultivated, and the cultured upper liquid is further cultivated by repeatedly removing to a new container.
In regards to treatment methods for cGVHD, U.S. Pat. No. 6,544,506 presents a GVHD prevention and treatment method that has as its distinctive feature the removal of cytotoxic T lymphocytes by injecting non-alloreactive anti-cytotoxic lymphocytes in organ transplantation patients. U.S. Pat. No. 6,936,281 describes a GVHD treatment method using mesenchymal stem cells. U.S. Pat. No. 7,173,016 describes a GVHD treatment method that includes the step of injecting adenosine deaminase inhibitors. U.S. Pat. No. 6,328,960 describes a GVHD treatment method that has as its distinctive feature the injection of mesenchymal stem cells in an amount that can lessen the immunological reaction of effector cells against the antigens in the target organ transplantation patient in order to lessen the effector cell's immunological reaction against the alloantigen in the target transplantation patient. U.S. Pat. No. 6,368,636 describes a method of lessening the immunological reaction caused by effector cells, which includes the step of contacting the effector cells with the upper liquid of the mesenchymal stem cells in an injection amount that can reduce the immunological reaction against alloantigen.
Until now, there is no report of a case in which cGVHD has been treated successfully using mesenchymal or marrow stem cells. Accordingly, the inventors herein through attempting to treat cGVHD completed this invention by verifying clinically that mesenchymal or marrow stem cells that were separated using a subfractionation culturing method effectively treats cGVHD.